1. What does nonclinical studies include?
Answer: Nonclinical experiments include pharmacology studies, general toxicity tests, toxicokinetics and nonclinical pharmacokinetics experiments, reproductive toxicity tests, genotoxicity tests, carcinogenicity, and other tests (phototoxicity, dependence, immunotoxicity), etc. In cases of combination therapy, nonclinical safety studies of combination therapy may also be conducted. For certain high proportion (greater than 10% of the total systemic exposure and the exposure level in animal studies is lower than human exposure), it may be required to conduct nonclinical safety studies of metabolites.
2. What is the purpose of nonclinical safety evaluation?
Answer: The purpose of nonclinical safety evaluation generally includes elucidating toxicity reaction characteristics related to target organs, dose dependency, exposure relationship, and potential reversibility. This information can be used to estimate the safe starting dose and dose range for human clinical trials (as mentioned earlier, NOAEL needs to be obtained in toxicology experiments for calculating MRSD, the maximum recommended starting dose), and to determine clinical monitoring indicators for potential adverse reactions.
3. What nonclinical studies are required for first-in-human trials?
Answer: For this question, it is first necessary to clarify the type of drug. ICH M3 (R2) is applicable to common situations in drug development and provides general guidance principles. For biologics, nonclinical studies should be conducted according to the requirements of ICH S6.
⑴Pharmacokinetics: Before IND submission, data on the metabolism of the investigational product in different species and in vitro metabolism data in humans, as well as plasma protein binding rate, are sufficient. Information on absorption, distribution, in vivo metabolism, and excretion should be completed before large-scale clinical trials (usually phase 3). PK absorption experiments are generally conducted in multiple species before IND submission and PCC determination to better understand the PK characteristics of the drug in different species, although it is not required here, which is a bit strange.
⑵Safety Pharmacology: Evaluation of the effects on cardiovascular, central nervous, and respiratory systems should be completed before IND submission, which is a combination of safety pharmacology standard tests. Supplementary safety pharmacology experiments may be conducted during clinical trials if necessary.
⑶Repeat Dose Toxicity: Generally, a short-term chronic toxicity test is arranged based on the early dosing scheme of IND. If the dosing period needs to be extended later, a longer-term chronic toxicity test is required to support it.
⑷Genotoxicity: With relatively low cost, genotoxicity test combination can be completed before IND submission.
⑸Carcinogenicity: Carcinogenicity testing should be completed before drug approval.
⑹Reproductive Toxicity: Observations and evaluations of the reproductive system should be included in repeat dose toxicity tests before IND submission to preliminarily assess reproductive toxicity, with strict contraception in clinical trials being sufficient. Phase I/II reproductive toxicity studies should be completed before large-scale clinical trials, and Phase III reproductive toxicity studies should be completed before approval.
⑺Phototoxicity: Phototoxicity assessment should be completed before IND submission. If the molar absorptivity is greater than 1000 M-1cm-1, there is considered to be a risk of phototoxicity, and phototoxicity testing should be conducted before large-scale clinical trials.
4. How should doses be set in toxicology tests? How should the highest dose be selected?
Answer: Referring to the "Technical Guidance Principles for Drug Repeated Dose Toxicity Tests," "In principle, high doses should induce obvious toxic reactions in animals, low doses should be equivalent to or higher than the effective dose in animals or clinical use, and middle doses should be set between high doses and low doses based on the toxic mechanism and characteristics to examine the dose-response relationship of toxicity."
In practice, considering the expensive cost of toxicology tests, conducting a dose range-finding test (DRF) with a few non-human primates is the most cost-effective and labor-saving method. The dose for DRF tests should also be further set based on preliminary safety studies conducted during early PCC determination and pharmacological studies. Many institutions now also conduct acute toxicity tests alongside DRF tests, which are non-GLP tests, as mentioned in M3 (R2), following the 3R principle to reduce the use of animals.
For low, medium, and high dose groups, a spacing of 2-3 times is generally used. In formal toxicology tests, the ideal situation is that the high-dose group can demonstrate obvious toxicity, and the low or medium dose groups can be used as NOAEL.
In addition, for the selection of the highest dose, M3 (R2) also provides standard answers. In simple terms, the highest dose in toxicology should either be the maximum tolerated dose (MTD) or the maximum feasible dose (MFD, which can be understood as the maximum dose that animals can tolerate based on product and animal characteristics), or it should achieve exposure saturation, or the exposure level should be sufficiently high (reaching 50 times the intended clinical dose). In general, 1000 mg/kg/day is considered sufficient unless the intended clinical dose reaches 1g/day, and the exposure level of 1000 mg/kg does not reach 10 times the clinical exposure level, in which case you need to further increase the dose to 10 times the clinical exposure level, or 2000 mg/kg/day, or MFD.
5. Which studies in nonclinical safety research need to follow GLP, and which ones do not?
Answer: Pharmacological and pharmacokinetic studies do not need to follow GLP, while safety pharmacology, genotoxicity, reproductive toxicity, and repeat dose toxicity studies need to follow GLP.
6. How long is the duration of nonclinical repeat dose toxicity studies?
Answer: The duration requirements for supporting clinical trial conduct and market approval are different and need to be specific to the dosing period of clinical trials. For example, for patients with multiple sclerosis, the minimum planned clinical trial period is two years. Therefore, when enrolling multiple sclerosis patients in clinical trials, a 6-month rodent and 9-month non-rodent chronic toxicity study is required, which is sufficient to support market approval. For non-oncology drugs, a 4-week chronic toxicity study is generally used to support a dosing period not exceeding 4 weeks for first-in-human clinical trials.
7. How to choose experimental animals for nonclinical safety evaluation?
Answer: Generally, two types of animals are used, rodents with rats being the preferred choice, and non-rodents with Beagle dogs being the preferred choice. Of course, specific product-specific analysis principles should also be followed. For small molecules, the selection of species should mainly consider the metabolic characteristics of the investigational product in animals that are similar to humans. Reference can be made to the "Analysis and Identification Test of Metabolites of Investigational Products in CD-1 Mice, SD Rats, Beagle Dogs, Cynomolgus Monkeys, and Human Liver Microsomes", selecting species with metabolic characteristics similar to humans, that is, species with similar amounts and types of metabolites to humans. Additionally, the selected species should be sensitive to the investigational product. Based on past experience, for a small molecule drug, its metabolic characteristics in Beagle dogs may be similar to humans but with low exposure.
